Dual Role of the RIC-3 Protein in Trafficking of Serotonin and Nicotinic Acetylcholine Receptors

Abstract

The ric-3 gene is required for maturation of nicotinic acetylcholine receptors in Caenorhabditis elegans. The human homolog of RIC-3, hRIC-3, enhances expression of ␣7 nicotinic receptors in Xenopus laevis oocytes, whereas it totally abolishes expression of ␣4␤2 nicotinic and 5-HT 3 serotonergic receptors. Both the N-terminal region of hRIC-3, which contains two transmembrane segments, and the C-terminal region are needed for these differential effects. hRIC-3 inhibits receptor expression by hindering export of mature receptors to the cell membrane. By using chimeric proteins made of ␣7 and 5-HT 3 receptors, we have shown that the presence of an extracellular isoleucine close to the first transmembrane receptor fragment is responsible for the transport arrest induced by hRIC-3. Enhancement of ␣7 receptor expression occurs, at least, at two levels: by increasing the number of mature receptors and facilitating its transport to the membrane. Certain amino acids of a putative amphipathic helix present at the large cytoplasmic region of the ␣7 subunit are required for these actions. Therefore, hRIC-3 can act as a specific regulator of receptor expression at different levels.

Ric-3 chaperone-mediated stable cell-surface expression of the neuronal α7 nicotinic acetylcholine receptor in mammalian cells

“…The levels of cell-surface α7 AChRs in this new cell line increased from less than 1% to 20% (Figure 1). On the basis of previous reports [13,14,16,17] , we can surmise that Ric-3 facilitates the correct folding and assembly of α7 subunits at the ER. When the parental SHE-P1-hα7 cells were labeled with fluorescent αBTX, a punctate distribution reminiscent of the ER staining was observed throughout the cell ( Figure 1B).…”

“…There is evidence that AChR folding, assembly and trafficking are influenced by several chaperone proteins. Recent studies have shown that co-expression of α7 AChR and the chaperone RIC-3 facilitates the formation of functional homomeric AChRs in otherwise non-permissive cell types [13,14,16,17] . RIC-3 was originally identified in the nematode Caenorhabditis elegans as a protein encoded by the gene ric-3 (resistance to inhibitors of cholinesterase) and has subsequently been cloned and characterized for mammalian and insect species.…”

“…Ric-3, first discovered in Caenorhabditis elegans, is an endoplasmic reticulum-resident protein present in cells that endogenously express α7 AChR [13] . Ric-3 has been shown to increase α7 AChR heterologous expression in oocytes from Xenopus laevis and in mammalian cell lines such as HEK-293 [13][14][15][16][17] . According to these authors, Ric-3 is needed to attain the correct folding of the α7 AChR, and therefore to attain functional expression.…”

Muscle-like nicotinic receptor accessory molecules in sensory hair cells of the inner ear

“…Recently, the human homolog RIC-3 has been shown to bind to α7 nAChRs and enhances their functional expression in Xenopus Laevis oocytes and mammalian cells (Castillo et al, 2005;Williams et al, 2005). However, the precise mechanisms by which the hRIC-3 protein exerts its effects on nicotinic receptors are not known.…”

A review of experimental techniques used for the heterologous expression of nicotinic acetylcholine receptors

“…As has been discussed elsewhere [95,287], severe difficulties have been encountered in obtaining functional expression of recombinant α7 nAChRs in several cultured mammalian cell lines [291][292][293][294][295][296]. Recent studies have revealed that coexpression of α7 with RIC-3 in such non-permissive cells facilitates appropriate folding and functional expression of α7 nAChRs [290,297,298]. In addition, it has been reported that co-expression of α4β2 nAChRs with UNCL, a mammalian homologue of the C. elegans transmembrane protein UNC-50, results in increased nAChR functional expression [299], although it has been suggested that this may be due to an RNA-binding …”